Name: a model of chronic nutrient infusion in the rat
Uploaded: 2013-08-14T12:00:00
Description: Watch this Scientific Journal Video about A Model of Chronic Nutrient Infusion in the Rat at JoVE.com

This work was supported by the National Institutes of Health (R01DK58096 to Vincent Poitout). Vincent Poitout holds the Canada Research Chair in Diabetes and Pancreatic Beta-cell Function. Bader Zarrouki received post-doctoral fellowships from Merck and Eli Lilly. Ghislaine Font&eacute;s was supported by a post-doctoral fellowship from the Canadian Diabetes Association.

Overview: The procedure consists of catheterizing the right jugular vein and left carotid artery under general anesthesia; allowing a 7-day recuperation period; connecting the catheters to the pumps using a swivel and counterweight system that enables the animal to move freely in the cage; and infusing glucose and/or Intralipid (a soybean oil emulsion which generates a mixture of approximately 80% unsaturated/20% saturated fatty acids when infused with heparin 9) for 72 hr.
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<ol>
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C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=High+glucose+stimulates+early+response+gene+c-Myc+expression+in+rat+pancreatic+beta+cells.">High glucose stimulates early response gene c-Myc expression in rat pancreatic beta cells.</a> The Journal of Biological Chemistry. 276, 35375-35381 (2001).</li><li>Tang, C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Glucose-induced+beta+cell+dysfunction+in+vivo+in+rats:+link+between+oxidative+stress+and+endoplasmic+reticulum+stress.">Glucose-induced beta cell dysfunction in vivo in rats: link between oxidative stress and endoplasmic reticulum stress.</a> Diabetologia. 55, 1366-1379 (2012).</li><li>Alonso, L. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Glucose+infusion+in+mice:+a+new+model+to+induce+beta-cell+replication.">Glucose infusion in mice: a new model to induce beta-cell replication.</a> Diabetes. 56, 1792-1801 (2007).</li><li>Magnan, C., Gilbert, M., Kahn, B. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Chronic+free+fatty+acid+infusion+in+rats+results+in+insulin+resistance+but+no+alteration+in+insulin-responsive+glucose+transporter+levels+in+skeletal+muscle.">Chronic free fatty acid infusion in rats results in insulin resistance but no alteration in insulin-responsive glucose transporter levels in skeletal muscle.</a> Lipids. 31, 1141-1149 (1996).</li><li>Goh, T. T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Lipid-induced+beta-cell+dysfunction+in+vivo+in+models+of+progressive+beta-cell+failure.">Lipid-induced beta-cell dysfunction in vivo in models of progressive beta-cell failure.</a> Am. J. Physiol. Endocrinol. Metab. 292, 549-560 (2007).</li><li>Pascoe, J., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Free+fatty+acids+block+glucose-induced+beta-cell+proliferation+in+mice+by+inducing+cell+cycle+inhibitors+p16+and+p18.">Free fatty acids block glucose-induced beta-cell proliferation in mice by inducing cell cycle inhibitors p16 and p18.</a> Diabetes. 61, 632-641 (2012).</li><li>Bell, C. G., Walley, A. J., Froguel, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+genetics+of+human+obesity.">The genetics of human obesity.</a> Nature Reviews. Genetics. 6, 221-234 (2005).</li><li>Fontes, G., Hagman, D. K., Latour, M. G., Semache, M., Poitout, V. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Lack+of+preservation+of+insulin+gene+expression+by+a+glucagon-like+peptide+1+agonist+or+a+dipeptidyl+peptidase+4+inhibitor+in+an+in+vivo+model+of+glucolipotoxicity.">Lack of preservation of insulin gene expression by a glucagon-like peptide 1 agonist or a dipeptidyl peptidase 4 inhibitor in an in vivo model of glucolipotoxicity.</a> Diabetes Res. Clin. Pract. 87, 322-328 (2010).</li><li>Crawford, P. A., Schaffer, J. 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Metab Care. 13, 382-390 (2010).</li><li>Cusi, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Role+of+obesity+and+lipotoxicity+in+the+development+of+nonalcoholic+steatohepatitis:+pathophysiology+and+clinical+implications.">Role of obesity and lipotoxicity in the development of nonalcoholic steatohepatitis: pathophysiology and clinical implications.</a> Gastroenterology. 142, 711-725 (2012).</li></ol>

Although a number of previous studies have employed chronic infusions of glucose (e.g. 10-15) or lipids (e.g. 16,17) in rodents, to our knowledge the combined infusion of both fuels has only been reported in mice 18. The chronic infusion model presented here offers several advantages to study the effects on nutrient excess on a variety of biological functions in rats. First, it does not involve genetically obese rodents, and since common obesity in humans is polyge...

Chronically elevated levels of glucose and lipids in the circulation have been proposed to contribute to the pathogenesis of type 2 diabetes by altering the function of several organs implicated in the maintenance of glucose homeostasis including, but not limited to, the pancreatic beta-cell (reviewed in 1). The glucotoxicity hypothesis posits that chronic hyperglycemia aggravates the beta-cell defect which gave rise to hyperglycemia in the first place, thus creating a vicious cycle and contributing to the deterioration of glucose control in type 2 diabetes patients. Likewise, the glucolipotoxicity hypothesis proposes that concomitant elevations of glucose and lipid levels, as often observed in type 2 diabetes, are detrimental to the beta cell.
	Deciphering the cellular and molecular mechanisms of the deleterious effects of chronically elevated nutrients on pancreatic beta-cell function is key to the understanding of the pathogenesis of type 2 diabetes 1. To that end, a large number of studies have examined the mechanisms of chronic nutrient excess ex vivo in isolated islets of Langerhans or in vitro in clonal, insulin-secreting cell lines. However, translation of the findings obtained in these model systems to the whole organism is complex, in particular because the concentrations of fatty acids used in cultured cells or islets rarely match the circulating levels in the vicinity of the beta cells in vivo 2. On the other hand, the mechanisms of beta-cell failure in response to nutrient excess have been examined in rodent models of diabetes, as exemplified by the Zucker Diabetic Fatty rat 3,4, the gerbil Psammomys obesus 5 and the high-fat diet-fed mouse 6. These models, however, are characterized by intrinsic metabolic abnormalities and are not easily amenable to manipulations of blood glucose and/or lipid levels in a more controlled and less chronic setting. To be able to alter circulating nutrient levels in a timeframe of days in otherwise normal animals, we have developed a chronic infusion model in normal rats which enables us to examine the effects of lipids and glucose, alone or in combination, on physiological parameters and function 7,8.

<table border="1">
		<tbody>
		 <tr>
			<td>Name of Reagent/Material</td>
			<td>Company</td>
			<td>Catalog Number</td>
			<td>Comments</td>
		 </tr>
		 <tr>
			<td>Saline 0.9%</td>
			<td>BD</td>
			<td>JB1324</td>
			<td></td>
		 </tr>
		 <tr>
			<td>Dextrose 70%</td>
			<td>McKesson</td>
			<td></td>
			<td></td>
		 </tr>
		 <tr>
			<td>Intralipid 20%</td>
			<td>Fresenius Kabi</td>
			<td>JB6023</td>
			<td></td>
		 </tr>
		 <tr>
			<td>Metricide (Glutaraldehyde 2.6%)</td>
			<td>Metrex</td>
			<td>11-1401</td>
			<td></td>
		 </tr>
		 <tr>
			<td>Heparin Sodium 10,000 USP u/ml</td>
			<td>PPC</td>
			<td></td>
			<td></td>
		 </tr>
		 <tr>
			<td>Carprofen</td>
			<td>Metacam</td>
			<td></td>
			<td></td>
		 </tr>
		 <tr>
			<td>Glycopyrrolate</td>
			<td>Sandoz</td>
			<td></td>
			<td></td>
		 </tr>
		 <tr>
			<td>Isoflurane</td>
			<td>Abbott</td>
			<td></td>
			<td></td>
		 </tr>
		 <tr>
			<td>Chlohexidine 2%</td>
			<td></td>
			<td></td>
			<td></td>
		 </tr>
		 <tr>
			<td>Alcohol 70%</td>
			<td></td>
			<td></td>
			<td></td>
		 </tr>
		 <tr>
			<td>Iodine</td>
			<td></td>
			<td></td>
			<td></td>
		 </tr>
		 <tr>
			<td>PE-50</td>
			<td>BD</td>
			<td>427411</td>
			<td></td>
		 </tr>
		 <tr>
			<td>CO-EX T22</td>
			<td>Instech Solomon</td>
			<td>BCOEX-T22</td>
			<td></td>
		 </tr>
		 <tr>
			<td>Connector 22G</td>
			<td>Instech Solomon</td>
			<td>SC22/15</td>
			<td></td>
		 </tr>
		 <tr>
			<td>Swivel 22G</td>
			<td>Instech Solomon</td>
			<td>375/22PS</td>
			<td></td>
		 </tr>
		 <tr>
			<td>Y-Connector 22G</td>
			<td>Instech Solomon</td>
			<td></td>
			<td></td>
		 </tr>
		 <tr>
			<td>Counterbalance and arm</td>
			<td>Instech Solomon</td>
			<td>CM375BP</td>
			<td></td>
		 </tr>
		 <tr>
			<td>23 G blunted needles</td>
			<td>Instech Solomon</td>
			<td>LS23</td>
			<td></td>
		 </tr>
		 <tr>
			<td>23 G canulation pins</td>
			<td>Instech Solomon</td>
			<td>SP23/12</td>
			<td></td>
		 </tr>
		 <tr>
			<td>Tethers (12 inch)</td>
			<td>Lomir</td>
			<td>RT12D</td>
			<td></td>
		 </tr>
		 <tr>
			<td>Infusion jackets </td>
			<td>Lomir</td>
			<td>RJ01, RJ02, RJ03, RJ04</td>
			<td></td>
		 </tr>
		 <tr>
			<td>(SM-XL)</td>
			<td></td>
		 </tr>
		 <tr>
			<td>Tether attachment piece</td>
			<td>Lomir</td>
			<td>RS T1</td>
			<td></td>
		 </tr>
		 <tr>
			<td>60 ml syringe</td>
			<td>BD</td>
			<td>309653</td>
			<td></td>
		 </tr>
		 <tr>
			<td>1 ml syringe</td>
			<td>BD</td>
			<td>309602</td>
			<td></td>
		 </tr>
		</tbody>
 </table>

a model of chronic nutrient infusion in the rat

Chronic exposure to excessive levels of nutrients is postulated to affect the function of several organs and tissues and to contribute to the development of the many complications associated with obesity and the metabolic syndrome, including type 2 diabetes. To study the mechanisms by which excessive levels of glucose and fatty acids affect the pancreatic beta-cell and the secretion of insulin, we have established a chronic nutrient infusion model in the rat. The procedure consists of catheterizing the right jugular vein and left carotid artery under general anesthesia; allowing a 7-day recuperation period; connecting the catheters to the pumps using a swivel and counterweight system that enables the animal to move freely in the cage; and infusing glucose and/or Intralipid (a soybean oil emulsion which generates a mixture of approximately 80% unsaturated/20% saturated fatty acids when infused with heparin) for 72 hr. This model offers several advantages, including the possibility to finely modulate the target levels of circulating glucose and fatty acids; the option to co-infuse pharmacological compounds; and the relatively short time frame as opposed to dietary models. It can be used to examine the mechanisms of nutrient-induced dysfunction in a variety of organs and to test the effectiveness of drugs in this context.

Out of a series of 42 rats which underwent surgery, 5 rats were lost during the post-operative period and 1 rat was lost during the infusion, representing an overall success rate of 86%. The average body weight of the 37 rats that were eventually infused was 608&plusmn;5 g before surgery and 588&plusmn;6 g at the initiation of the infusion (mean&plusmn;SE; n=37; P&lt;0.0001 by paired t-test). The following representative results were obtained in 2 infusion groups: Saline (SAL), and Glucose + Intralipid (GLU+IL). ...

Watch this Scientific Journal Video about A Model of Chronic Nutrient Infusion in the Rat at JoVE.com

A Model of Chronic Nutrient Infusion in the Rat

A protocol for chronic infusions of glucose and Intralipid in rats is described. This model can be used to study the impact of nutrient excess on organ function and physiological parameters.

Chronic exposure to excessive levels of nutrients is postulated to  affect the function of several organs and tissues and to contribute to the  ...

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